Amphotericin B resistance in Lomentospora prolificans is associated with a soluble cell wall component

Lomentospora prolificans is a pathogenic filamentous fungus that causes disease primarily in people with severely compromised immune systems. It is pan-resistant to antifungal drugs, but the mechanism of its resistance to amphotericin B (AMB) is unknown. In this study, our goal was to investigate the mechanism of resistance to AMB of L. prolificans. The AMB susceptibility of L. prolificans protoplasts were measured using broth microdilution. L. prolificans, either intact, homogenized, or fractionated, was incubated with AMB in broth. The same activity was carried out with Aspergillus fumigatus as a control. These broths were then used to prepare microdilution plates with Saccharomyces cerevisiae to determine the activity of the conditioned AMB. AMB was 16-fold more effective at inhibiting the growth of L. prolificans protoplasts than conidia, but only 2-fold more effective against A. fumigatus protoplasts than conidia. Incubation of L. prolificans hyphae with AMB in media diminished drug activity to a much greater extent than A. fumigatus, with an 8-fold greater fungal mass of the latter required to achieve the effect of the former. Homogenization and fractionation of L. prolificans revealed that the factor inhibiting AMB activity was soluble with a mass >100 kDa. DNase, trypsin, proteinase K, amyloglucosidase, SDS, and 0.22 μm filtration had no effect on the AMB resistance factor, while treatment with urea and acetonitrile inactivated it. In summary, we report a different mechanism for AMB resistance for L. prolificans based on the existence of a substance residing in the cell wall that can eliminate the antifungal activity of AMB.